Marked elevation in cortical urate and xanthine oxidoreductase activity in experimental bacterial meningitis

Experimental bacterial meningitis due to Streptococcus pneumoniae in infant rats was associated with a time-dependent increase in CSF and cortical urate that was approximately 30-fold elevated at 22 h after infection compared to baseline. This increase was mirrored by a 20-fold rise in cortical xanthine oxidoreductase activity. The relative proportion of the oxidant-producing xanthine oxidase to total activity did not increase, however. Blood plasma levels of urate also increased during infection, but part of this was as a consequence of dehydration, as reflected by elevated ascorbate concentrations in the plasma. Administration of the radical scavenger alpha-phenyl-tert-butyl nitrone, previously shown to be neuroprotective in the present model, did not significantly affect either xanthine dehydrogenase or xanthine oxidase activity, and increased even further cortical accumulation of urate. Treatment with the xanthine oxidoreductase inhibitor allopurinol inhibited CSF urate levels earlier than those in blood plasma, supporting the notion that urate was produced within the brain. However, this treatment did not prevent the loss of ascorbate and reduced glutathione in the cortex and CSF. Together with data from the literature, the results strongly suggest that xanthine oxidase is not a major cause of oxidative stress in bacterial meningitis and that urate formation due to induction of xanthine oxidoreductase in the brain may in fact represent a protective response.

Anteroposterior pelvic radiographs to assess acetabular retroversion: high validity of the "cross-over-sign"

Acetabular retroversion has been proposed to contribute to the development of osteoarthritis of the hip. For the diagnosis of this condition, conventional AP pelvic radiographs may represent a reliable, easily available diagnostic modality as they can be obtained with a reproducible technique allowing the anterior and posterior acetabular rims to be visible for assessment. This study was designed to: (i) determine cranial, central, and caudal anatomic acetabular version (AV) from cadaveric specimens; (ii) establish the validity and reliability of the radiographic measurements of central acetabular anteversion; and (iii) determine the validity and reliability of the radiographic "cross-over-sign" to detect acetabular retroversion. Using 43 desiccated pelvises (86 acetabuli) the anatomic AVs were measured at three different transverse planes (cranially, centrally, and caudally). From these pelvises, standardized AP pelvic radiographs were obtained. To directly measure central AV, a modified radiographic method is introduced for the use of AP pelvic radiographs. The validity and reliability of this radiographic method and of the radiographic cross-over-sign to detect cranial acetabular retroversion were determined. The mean central and caudal anatomic AVs were approximately 20 degrees , and the mean cranial AV was 8 degrees . Cranial retroversion (AV < 0 degrees ) was present in 19 of 86 hips (22%). A linear correlation was found between the central and cranial AV. Below 10 degrees of central AV, all acetabuli were cranially retroverted. Between 10 degrees and 20 degrees , 30% of the acetabuli were cranially retroverted, and above 20 degrees , only 1 of 45 acetabuli was cranially retroverted. The radiographic measurement of the central AV (20.3 +/- 6.5 degrees ) correlated strongly with the anatomic AV (20.1 +/- 6.4 degrees ). The sensitivity of the cross-over-sign to detect a cranial acetabular anteversion of less than 4 degrees was 96%, its specificity 95%, and the positive predictive and negative predictive values 90% and 98%, respectively. Both the modified radiographic anteversion measurements and the cross-over-sign demonstrated substantial inter- and intraobserver reliability. Retroversion is almost exclusively a problem of the cranial acetabulum. The cranial AV is on average 12 degrees lower than the central AV, with the latter directly measurable from AP pelvic radiographs. A central AV of less than 10 degrees was associated with cranial retroversion. The presence of a positive cross-over-sign is a highly reliable indicator of cranial AV of <4 degrees.

EpCAM-targeted delivery of nanocomplexed siRNA to tumor cells with designed ankyrin repeat proteins

Specific delivery to tumors and efficient cellular uptake of nucleic acids remain major challenges for gene-targeted cancer therapies. Here we report the use of a designed ankyrin repeat protein (DARPin) specific for the epithelial cell adhesion molecule (EpCAM) as a carrier for small interfering RNA (siRNA) complementary to the bcl-2 mRNA. For charge complexation of the siRNA, the DARPin was fused to a truncated human protamine-1 sequence. To increase the cell binding affinity and the amount of siRNA delivered into cells, DARPin dimers were generated and used as fusion proteins with protamine. All proteins expressed well in Escherichia coli in soluble form, yet, to remove tightly bound bacterial nucleic acids, they were purified under denaturing conditions by immobilized metal ion affinity chromatography, followed by refolding. The fusion proteins were capable of complexing four to five siRNA molecules per protamine, and fully retained the binding specificity for EpCAM as shown on MCF-7 breast carcinoma cells. In contrast to unspecific LipofectAMINE transfection, down-regulation of antiapoptotic bcl-2 using fusion protein complexed siRNA was strictly dependent on EpCAM binding and internalization. Inhibition of bcl-2 expression facilitated tumor cell apoptosis as shown by increased sensitivity to the anticancer agent doxorubicin.

High-resolution summer temperature reconstruction from Lake Silvaplana based on in-situ reflectance spectroscopy

Annually laminated (varved) sediments of proglacial Lake Silvaplana (46 ̊27’N, 9 ̊48’E, 1791 m a.s.l., Engadine, eastern Swiss Alps) provide an excellent archive for quantitative high-resolution (seasonal – annual) reconstruction of high- and lowfrequency climate signals back to AD 1580. The chronology of the core is based on varve counting, Cs-137, Pb-210 and event stratigraphy. In this study we present a reconstruction based on in-situ reflectance spectroscopy. In situ reflectance spectroscopy is known as a cost- and time-effective non destructtive method for semi-quantitative analysis of pigments (e.g., chlorines and carotenoids) and of lithoclastic sediment fractions. Reflectance-dependent absorption (RDA) was measured with a Gretac Macbeth spectrolino at 2 mm resolution. The spectral coverage ranges from 380 nm to 730 nm at 10 nm band resolution. In proglacial Lake Silvaplana, 99% of the sediment is lithoclastic prior to AD 1950. Therefore, we concentrate on absorption features that are characteristic for lithoclastic sediment fractions. In Lake Silvaplana, two significant correlations that are stable in time were found between RDA typical for lithoclastics and meteorological data: (1) the time series R 570 /R 630 (ratio between RDA at 570 nm and 630 nm) of varves in Lake Silvaplana and May to October temperatures at nearby station of Sils correlate highly significantly (calibration period AD 1864 – 1951, r = 0.74, p < 0.01 for 5ptsmoothed series; RMSE is 0.28 ̊C, RE = 0.41 and CE = 0.38), and (2) the minimum reflectance within the 690nm band (min690) data correlate with May to October (calibration period AD 1864 – 1951, r = 0.68, p < 0.01 for 5pt-smoothed series; RMSE = 0.22 ̊C, RE = 0.5, CE = 0.31). Both proxy series (min690nm and R 570 /R 630 values) are internally highly consistent (r = 0.8, p < 0.001). In proglacial Lake Silvaplana the largest amount of sediment is transported by glacial meltwater. The melting season spans approximately from May to October, which gives us a good understanding of the geophysical processes explaining the correlations between lithoclastic proxies and the meteorological data. The reconstructions were extended back to AD 1580 and show a broad corresponddence with fully independent reconstructions from tree rings and documentary data.